Nylon-6-Coated Doxorubicin-Loaded Magnetic Nanoparticles and Nanocapsules for Cancer Treatment

Round 1

Reviewer 1 Report

The manuscript by Kovrigina et al. describes a method for loading doxorubicin into magnetic nanocomposites or nanocapsules. Ultimately, the authors concluded that oleic-acid NC2_DOX had excellent prospects for cancer treatments. Based on the citations, the authors have done significant work in this area, and the paper is very well received. A few revisions for this paper are listed below:

- Please add the UV-Vis calibration curves for DOX adsorption/release studies in the supplemental section. Also, indicate which wavenumber was used for absorbance readings. 

- In the DOX loading and Release sections (3.6 and 3.7), please indicate centrifugation speed and time so that other researchers can use this information in other studies. 

- Cell viability studies were performed with DOX. However, the novel nanocapsules (NC1 and NC2) were not evaluated (i.e., without DOX). Therefore, it is critical to demonstrate the cell viability results on the capsules alone. I will highly suggest this addition to make the paper highly influential and to re-confirm the biocompatibility of Nylon in the form of a nanocapsule. 

- Figure 5 shows DOX release results. This figure is crucial to the study. However, the loading was different for each NC or MNC. Please try to change (or add a supplemental figure) the DOX release figure by using a normalized plot for the individual DOX loading of each NC or MNC. A normalized percent release plot will help discuss and present the results. 

- Ideally, drug release as a function of time for the most promising compound - at least  NC2 - should be provided. 

- Please try to discuss why NC2 is better than NC1. The only difference is the OA, hence why oleic acid is an essential factor in the release of DOX. 

Author Response

Thank you for the valuable suggestions and comments. We have carefully examined the comments and suggestions and revised the manuscript accordingly. We presented the word file with track changes. Please find as follows the responses to the comments. Please note that all the comments are bold-faced, and the authors' reply follows immediately below the comments.

- Please add the UV-Vis calibration curves for DOX adsorption/release studies in the supplemental section. Also, indicate which wavenumber was used for absorbance readings. 

Thank you for the suggestion. We added the information and the wavenumber in supplementary materials (Figure S1) and section 3.2.

- In the DOX loading and Release sections (3.6 and 3.7), please indicate centrifugation speed and time so that other researchers can use this information in other studies. 

Thank you for the suggestion. We have added the data in these sections.

- Cell viability studies were performed with DOX. However, the novel nanocapsules (NC1 and NC2) were not evaluated (i.e., without DOX). Therefore, it is critical to demonstrate the cell viability results on the capsules alone. I will highly suggest this addition to make the paper highly influential and to re-confirm the biocompatibility of Nylon in the form of a nanocapsule. 

Thank you for your valuable comments. We inserted a new cell viability Figure 7 in section 2.5.

- Figure 5 shows DOX release results. This figure is crucial to the study. However, the loading was different for each NC or MNC. Please try to change (or add a supplemental figure) the DOX release figure by using a normalized plot for the individual DOX loading of each NC or MNC. A normalized percent release plot will help discuss and present the results. 

Thank you for the suggestion. However, at the moment, we have Table 4, which contains these data. We understand that the DOX release is essential for the study and enlarged the discussion of these data.

- Ideally, drug release as a function of time for the most promising compound - at least NC2 - should be provided. 

Thank you for the suggestion. We have inserted Figures S3 and S4.

- Please try to discuss why NC2 is better than NC1. The only difference is the OA, hence why oleic acid is an essential factor in the release of DOX. 

Thank you for the suggestion. We have tried to improve the text throughout the paper (see sections 2.3 and 2.4).

Reviewer 2 Report

The current work focuses on Nylon-6 Coated Doxorubicin-loaded Magnetic Nanoparticles and Nanocapsules for Cancer Treatment. The author’s some effort into the manuscript but major issues should be addressed. 

Abstract 

- Line 11-16, general information, rephrases the abstract section with the novelty of the current work and main outputs

Introduction

- The introduction is providing sufficient background and the most relevant references are included, but the novelty of this work is not highlighted and the author's contribution was unclear compared to other previous works.

Materials and equipment

-Some specific properties are required and should be inserted of used Nylon-6 to clear to the reader

- Line 212, ultrasonic bath, power?

-DOX loading was studied using UV-vis spectroscopy, at which wavelength? 

-No information about the type and model of used UV-vis spectroscopy

Results and Discussion 

-The discussion of TEM analysis is not informative, particle size distribution should be inserted

- Also more details are required on the discussion of zeta potential, why difference in charge? Which is more stable with high or low value?…

-The proposed nanocomposites according to a four-step procedure!! No confirmation in the text to support this claim. More analyses are required to confirm the proposed materials FTIR, XRD, and elemental analysis

Conclusions

Line 269, narrow size distribution!!! This claim is not confirmed in the text

Author Response

Thank you for the valuable suggestions and comments. We have carefully examined the comments and suggestions and revised the manuscript accordingly. We presented the word file with track changes. Please find as follows the responses to the comments. Please note that all the comments are bold-faced, and the authors' reply follows immediately below the comments.

Abstract Line 11-16, general information, rephrases the abstract section with the novelty of the current work and main outputs

Thank you for your valuable comments. We have changed the text.

Introduction The introduction is providing sufficient background and the most relevant references are included, but the novelty of this work is not highlighted and the author's contribution was unclear compared to other previous works.

Thank you for your valuable comments. We have changed the text

Materials and equipment

-Some specific properties are required and should be inserted of used Nylon-6 to clear to the reader

Thank you for the suggestion. However, there is a lot number of Nylon, and any related data may be found on the manufacturer (Sigma) website. We inserted some characterization data in section 3.1.

- Line 212, ultrasonic bath, power?

Thank you for your valuable comments. We have inserted the data in section 3.4.

-DOX loading was studied using UV-vis spectroscopy, at which wavelength? 

Thank you for your valuable comments. We have changed the text in sections 3.6 and 3.7. Wavelength 480 nm.

-No information about the type and model of used UV-vis spectroscopy

Thank you for your valuable comments. We have changed the text in section 3.2.

Electronic absorption spectra were recorded on a UV-2100 spectrometer (Shimadzu, Kyoto, Japan) or microplate reader Clariostar (BMG, Ortenberg, Germany). 

Results and Discussion 

-The discussion of TEM analysis is not informative, particle size distribution should be inserted

Thank you for the suggestion. We have tried to improve the text.

- Also more details are required on the discussion of zeta potential, why difference in charge? Which is more stable with high or low value?…

Thank you for the suggestion. We have tried to improve the text (see about charge section 2.1 and for zeta potential sections 2.1 and 2.2).

-The proposed nanocomposites according to a four-step procedure!! No confirmation in the text to support this claim. More analyses are required to confirm the proposed materials FTIR, XRD, and elemental analysis

The synthesis was carried out according to the procedure developed earlier by the authors [1,2] using widely used tetraethyl orthosilicate/3-aminopropyltriethoxysilane (TEOS/APTES) method [3–9] with subsequent 2,4,6-trichloro-1,3,5-triazine reaction. TEOS/APTES/2,4,6-trichloro-1,3,5-triazine procedure is commonly known and was previously published. In this way, the extensive characterization of composites is held only after the whole synthesis pathway. Moreover, it should be noted that the reaction with TEOS was done in a water/ethanol mixture, with APTES in ethanol, with 1,3,5-trichloro-2,4,6-triazine in acetonitrile, with nylon in trifluoroethanol/water mixture. In this way, if we will take the reaction solutions for DLS, different solvent results will occur. It is not possible to compare the results for such systems in different solvents due to the various water sphere or solvent resize effects. Moreover, the size of nanoparticles in water is important for biomedical applications. If we will take the amount of the nanoparticles for TEOS and APTES stages and resolve them in water, Si-OH and NH₂ groups will fully interact with water, yielding a big water sphere by DLS. Nanoparticles after the 1,3,5-trichloro-2,4,6-triazine stage will be surface active and will interact with water by chemical reaction. In this way, the DLS method, which is commonly used in our laboratory, is not suitable for such a comparison. We used the DLS method for such synthesis only for reproducibility of the synthesis procedure in this non-water solvent stage to stage. In this way, we respectfully ask the reviewer to evaluate the characterization of the final nanoparticles by TEM, DLS, and FTIR. We have inserted FTIR data with a discussion in section 2.2, which easily shows different layers' peaks.

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Conclusions Line 269, narrow size distribution!!! This claim is not confirmed in the text

Thank you for your valuable comments. We have revised the text.

Round 2

Reviewer 1 Report

all comments addressed

Author Response

Thank you for your valuable comments.

Reviewer 2 Report

Accept in present form

Author Response

Thank you for your valuable comments.